High grade serous ovarian cancer (HGSC) is the most common and deadliest form of ovarian cancer. Emerging evidence indicates that these tumors arise in the fallopian tube epithelium (FTE), and thus their presence in the ovary represents the primary metastasis. Our preliminary data identified that xenografting in close proximity to the ovary contributes to the aggressiveness of the disease. After ovarian colonization, tumor cells invade the peritoneal organs, primarily the omentum. We hypothesize that the biological problem of primary and secondary HGSC metastasis is partially mediated by chemical communication between the cancer cells and the metastatic organ. Our proposal seeks to define metabolites and biomolecules that drive the metastasis of fallopian tube derived high grade serous cancers to the ovary and the omentum. To this end, our teams optimized a 3D co-culture of the ovary and fallopian tube derived tumor models and adapted this to imaging mass spectrometry technology to identify the metabolomics-driven communication that occurs during primary colonization of the ovary and during secondary metastasis to the omentum. Using this emerging technology, we identified several metabolites that enhanced high grade serous tumor migration, invasion, and adhesion to the ovary. The focus of Aim 1 is to uncover the mechanisms allowing FTE tumorigenic cells to hijack NE produced by the ovary to increase their ability to invade and adhere to the ovary during primary metastasis. Aim 1 will define the signaling pathways mediated by NE during invasion and adhesion to the ovary and then confirm the importance of NE in vivo using both murine and human cell models derived from FTE. The key adrenergic receptor will be deleted using CRISPR to confirm the importance of this pathway in metastasis. Tumor bearing models will be treated with beta adrenergic receptor antagonists in an attempt to translate these findings for a new strategy to block ovarian colonization. The focus of Aim 2 is on the identification and characterization of a newly identified protein that is secreted from tumorigenic fallopian tube cells and is responsible for the production of ovarian norepinephrine driving tumor cell invasion and adhesion. We will use proteomics to confirm the identity of the secreted protein, followed by genetic deletion of the protein from FTE models to study the role in ovarian colonization. Aim 3 will build upon our existing technology of 3D organ and tumor cell communication models and expand into secondary metastasis. We have now optimized our technology for co-culture of the omentum together with tumor cell models and have an inventory of metabolites, which are unique and did not include norepinephrine. Instead a novel metabolite found to be produced in significantly more abundance when tumor cells were grown with the omentum corresponded to folate, the ligand for the folic acid receptor that is overexpressed in the tumor cells. Taken together, our innovative experimental approach will yield new pathways and targets to mitigate primary metastasis of high grade serous cancer to the ovary.